A general feature that appears to be emerging for the structure of biological membranes is the existence of an asymmetrical phospholipid distribution between the membrane surfaces. Of fundamental importance to the discovery of such asymmetry are questions regarding its assembly and maintenance. Studies on asymmetry of membranes have been greatly facilitated by the ability to produce inside-out membrane vesicles. We propose to develop a technique which has general applicability to the study of lipid distributions in membranes by membrane inversion using positively charged glass beads. We will demonstrate that phospholipases and chemical labels can be used to determine the distribution of phospholipids in red cells attached to these beads. Subsequently, we will measure the distribution, biosynthetic assembly and possible transbilayer movement of the lipids in rat liver microsomes using inverted bead bound microsomal membranes. From this study we will be able to ascertain the mechanism(s) responsible for the assembly of lipid asymmetry in a biological membrane.